68 research outputs found
Improved detection of terminal transferase (TdT): The use of detergents on glutaraldehyde-fixed non-dehydrated cells prevents denaturation and diffusion artifacts.
TdT as an intranuclear enzyme mainly of immature lymphoid cells is commonly determined immunologically using air-dried cell smears fixed with methanol. Both cell dehydration and alcohol fixation were found here to denature TdT and surface antigens. This could be prevented by using non-dehydrated cells bound electrostatically to poly-L-lysine-coated slides, fixed minimally with glutaraldehyde and rendered permeable to antibodies by the non-ionic detergent Brij 56. Crosslinking glutaraldehyde in addition prevented diffusion of TdT to extranuclear sites. By avoiding artifacts of denaturation and diffusion, a higher sensitivity in the detection of TdT was achieved despite considerably lower quantities of antibody
Immunoglobulin expression on theta-positive leukemic cells in AKR/J mice.
The expression of surface immunoglobulin was studied on cells of theta-positive AKR leukemia during repeated weekly passages of ascitic cells. It could be shown that the percentage of Ig-positive cells varied considerably (0-25%) and that this was accompanied by variations in the expression of theta-antigen on the cell surface. This was interpreted as repression and derepression of genes coding for surface antigen possibly at the translational level
Effective avidin coating of erythrocytes by disulfide bond formation : Production of a stable rosetting agent for multiple applications.
A method is described for covalently coupling avidin to the membrane of glutaraldehyde-treated erythrocytes. The method is based on the use of the heterobifunctional coupling reagent N-succinimidyl-3-(2-pyridyldithio)propionate (SPDP) and appears to be more effective than other protein coupling techniques. The resulting avidin-coated red cells are very stable and can be stored for more than 1 year without aggregation or loss of biotin-binding activity. They can be used in various rosetting assays and were tested for T cell depletion or negative cell selection with five biotinylated monoclonal antibodies
Experimentelle Knochenmarkttransplantation (KMT). Neue Ansätze zur Überwindung transplantationsimmunologischer Komplikationen mittels Clq-Affiner Antikörper.
Antilymphocytic antibodies and marrow transplantation. XII. Suppression of graft-versus-host disease by T-cell-modulating and depleting antimouse CD3 antibody is most effective when preinjected in the marrow recipient.
A hamster antimouse CD3 monoclonal antibody (MoAb) opened the way to experimental studies on the suppression of allograft rejection and cytokine- related morbidity after treatment with antibodies modulating the CD3/T-cell receptor complex (CD3/TCR). Because earlier attempts to suppress graft- versus-host disease (GVHD) in patients by in vitro treatment of donor marrow with anti-CD3 MoAb had remained inconclusive, we used a rat IgG2b antimouse CD3 MoAb (17A2) with fewer side effects to analyze suppression of GVHD in the mouse model. Detailed phenotyping of blood, spleen, and lymphnode T cells after the injection of 400 μg 17A2 in C57BL/6 mice showed 60% CD3 downmodulation and 50% T-cell depletion for spleen cells. Injection of these spleen cells, together with bone marrow cells, in fully mismatched preirradiated CBA mice delayed GVHD by only 6 days. Ex vivo treatment of donor cells with 17A2 was not effective. In contrast, conditioning of marrow recipients with a single injection of 17A2 delayed 50% GVHD mortality by 100 days and prevented GVHD altogether after prolonged treatment, with survivors showing complete chimerism and specific transplantation tolerance. This difference in antibody effect contrasts with earlier experiences with nonmodulating but more strongly T-cell-depleting MoAbs of the same isotype, which prevent GVHD no matter whether applied in vitro or injected into donor or recipient mice. Our data indicate that CD3/TCR reexpression in marrow recipients with no circulating 17A2 is the reason why ex vivo donor cell treatment with anti-CD3 MoAb is comparatively ineffective. Our data, which allow separate evaluation of cell-depleting and cell-modulating antibody activity, help to explain previous clinical failure to suppress GVHD and provide evidence in favor of conditioning the marrow recipient with anti-CD3 MoAb as a therapeutic alternative
Coinjection of anti-interleukin-2 receptor antibody diminishes antiantibody formation during prolonged treatment with anti-CD3 antibody.
Poly-L-lysine as a coating agent for the attachment of active antibodies onto polyacrylic plastic beads.
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